Glycans perform varied and crucial functions in numerous cellular activities. The diverse roles of glycans are matched by their highly complex structures, which derive from differences in composition, branching, regio- and stereochemistry, and modification. This incomparable structural diversity is challenging to the structural analysis of glycans.
Recently, a joint research group led by Prof. QING Guangyan and Prof. LIANG Xinmiao from the Dalian Institute of Chemical Physics (DICP) of the Chinese Academy of Sciences (CAS) has developed a glycan identification method based on nanopore single-molecule sensing through a glycan derivatization strategy.
The study was published in Nature Communications on March 28.
Clear discrimination among four glycan isomers by single nanopore analysis, the glycans are labelled with a benzene tag (Image by Minmin Li)
Identifying and sequencing glycans using nanopore single-molecule techniques has sparked potential interests. However, it has achieved little progress over the past dozen years. Only a handful of cases that focused on either high molecular weight polysaccharides or some monosaccharides were reported.
For smaller but structurally more diverse glycans with greater biological significance, single molecule detection with nanopore has not yet been achieved, largely because fast passage of glycan through nanopore cannot be sensed because of the small size and weak affinity of glycan with nanopore.
To address the challenge, the researchers introduced a derivatization strategy by linking an aromatic-type tag group to small glycans via a high-efficiency and facile reductive amination reaction. The resulting tagged glycan was sensed with a wild-type aerolysin nanopore by presenting strong nanopore blockage signals.
The researchers obtained scatter plot based on blockage current and dwell time as the fingerprint map by processing the nanopore single-molecule blockage events. They identified different glycan isomers, glycans with varying lengths, and branched simple glycans.
Moreover, they revealed that multiple cation-π interactions between the aromatic tag of glycan with K238 residues of nanopore interface retarded the translocation of tagged glycan and contributed to the sensing.
"This study pushes the boundary of nanopore sensing beyond its traditional focus on nucleic acid and protein, and activates its power in the glycomics and glycoscience field, which might pave the way towards nanopore glycan sequencing", said Prof. QING.
This work was supported by the National Natural Science Foundation of China and the Innovation Grant of DICP.